tbuckley/cabot-search huggingface dataset:
TMET-23. GLIOBLASTOMA EXHIBIT SEX DIFFERENCES IN BRANCHED-CHAIN AMINO ACID METABOLISM
Authors: Jasmin Sponagel, Hannah Gass, Kevin Cho, Prakash Chinnaiyan, Gary Patti, Joshua Rubin, Joseph Ippolito
Journal: Neuro-Oncology (2022)

Abstract Glioblastoma (GBM) is the most common and aggressive primary brain tumor in adults. It is more prevalent in males and female patients exhibit a survival advantage. Understanding the molecular mechanisms that underlie those sex differences could support novel treatment strategies. Amino acids, such as glutamine and the branched-chain amino acids (leucine, isoleucine, valine; BCAAs), play important roles in GBM as they are involved in biosynthesis, energetics, redox homeostasis, and mTOR signaling. Using murine transformed astrocytes, we recently discovered that male cells are more dependent on glutamine and utilize more glutamine to replenish biosynthetic pathways, including BCAA regeneration. BCAA regeneration (i.e., re-amination of branched-chain ketoacids to BCAA) is catalyzed by branched-chain aminotransferases (BCATs) and requires glutamate as a nitrogen donor. Importantly, BCAA regeneration has been linked to cancer progression and poor outcome. Using stable isotope glutamine labeling we found that male transformed astrocytes exhibit greater nitrogen transfer from glutamate onto branched-chain ketoacids to regenerate BCAAs. In line with increased glutamate utilization for BCAA regeneration via BCAT, we found that male transformed astrocytes exhibit greater BCAT1 protein expression levels. Concordantly, male transformed astrocytes are more sensitive to BCAA deprivation compared to female cells, indicating that BCAA metabolism may play a sex-specific role in GBM. Using human GBM surgical specimens, we found greater BCAA metabolite abundance in male GBM. Importantly, our findings on sex differences in BCAA abundance and dependency in GBM are in line with previous reports that show that BCAA abundance and utilization differ between males and females under normal physiological conditions. Together, these data suggest that male GBM are more dependent on BCAAs and that BCAT1 may play unique roles in male and female GBM metabolism. Our data highlight that GBM metabolism differs between males and females and underline the importance of considering sex in metabolic targeting approaches.
IMMU-19. IDENTIFICATION OF TARGET ANTIGENS FOR CHIMERIC ANTIGEN RECEPTOR T- CELL THERAPY AGAINST GLIOBLASTOMA USING A MONOCLONAL ANTIBODY LIBRARY RAISED AGAINST PATIENT-DERIVED TUMOR SPHERES
Authors: Noriyuki Kijima, Tomoyoshi Nakagawa, Kana Hasegawa, Shunya Ikeda, Moto Yaga, Tansri Wibowo, Hideki Kuroda, Ryuichi Hirayama, Yoshiko Okita, Tetsuro Tachi, Naoki Kagawa, Yonehiro Kanemura, Naoki Hosen, Haruhiko Kishima
Journal: Neuro-Oncology (2022)

Abstract New therapies for GBM are urgently needed due to its poor prognosis and chimeric antigen receptor T (CAR-T) cell therapy is thought to be a promising strategy. To develop CAR-T cell therapy, cell surface targets that is highly specific for GBM cells are needed.Although extensive transcriptome analyses of GBM cells were performed, few transcripts highly specific for GBM cells have been identified. However, GBM cell-specific antigen epitopes formed by post-translational modifications of proteins may have been missed, and could still be discovered by thoroughly searching for cancer-specific monoclonal antibodies and characterizing the antigens they recognize. In this study, we applied this strategy to search for GBM-specific cell surface targets using patient derived tumor spheres. We identified two monoclonal antibodies E61 and A13 as those reacting with GBM cells but not with normal brain parenchymal cells. CAR-T cells derived from both monoclonal antibodies produced IL-2 and IFNɤ and exerted cytotoxicity to GBM cells by chromium 51 release assay. In addition, we identified B7-H3, which is frequently used for a CAR-T cell target against GBM, as the antigen recognized by B7-H3 by the expression cloning method. These results indicate that the strategy shown in this study is useful for identifying antigens that are expressed on patient-derived GBM cells and potentially useful as targets for CAR T cells.
IMMU-30. THE ROLE OF CD4 T CELLS IN ANTITUMOR IMMUNITY AGAINST BRAIN TUMORS
Authors: Andrew Coxon, Rupen Desai, Alexandra Livingstone, Dale Kobayashi, Gavin Dunn, Tanner Johanns
Journal: Neuro-Oncology (2022)

Abstract Glioblastoma is a primary brain tumor with a dismal prognosis. CD4 T cell responses to neoantigen vaccines or checkpoint blockade immunotherapy impact the tumor microenvironment. However, the function of CD4 T cells in glioblastoma remains understudied. Here we further characterize the role of CD4 T cells in antitumor immunity against the GL261 orthotopic model of glioblastoma. We studied the memory response generated during high-dose PD-L1 blockade mediated clearance of intracranial tumors. Mice that previously rejected GL261 retained protection against intracranial rechallenge after depletion of CD4 or CD8 T cells, but not both. Thus, these memory CD4 T cells are sufficient but not necessary for protection. Interestingly, in mice with a memory response generated during clearance of extracranial GL261, depletion of CD4 T cells abrogates protection against intracranial rechallenge, but depletion of CD8 T cells alone does not affect protection. Therefore, memory CD4 T cells generated with extracranial antigen challenge are necessary and sufficient for protection against intracranial rechallenge. In an analogous setting, we vaccinated mice against MHC class I neoantigens with concurrent checkpoint blockade immunotherapy to generate memory responses. Similar to the challenge-rechallenge model, mice challenged with GL261 and depleted of CD4 T cells lost protection from neoantigen vaccination and PD-L1 blockade. We then modified GL261 to express the C-terminal end of ovalbumin (GL261-OVAII), which contains the MHC class II immunodominant antigen (OVA323-339) but not the class I antigen (OVA257-264). Mice challenged intracranially with GL261-OVAII have a minimal survival benefit from PD-L1 blockade therapy. However, vaccination against OVA323-339 greatly enhances the protection from PD-L1 blockade. We determined by ELISPOT that CD4 T cells do not directly recognize GL261. Our data show the importance of the CD4 T cell subset in glioblastoma and the importance of targeting class II neoantigens to generate strong antitumor immunity.
EPCO-22. SINGLE CELL ANALYSIS TO UNDERSTAND SEX DIFFERENCES IN GBM OUTCOME
Authors: Lorida Llaci, Joshua Rubin, Rob Mitra
Journal: Neuro-Oncology (2022)

Abstract Males have a higher incidence and mortality in certain cancers compared to females. The same is true in glioblastoma (GBM) regardless of age, suggesting that cell-intrinsic factors rather than extrinsic factors such as circulating sex hormones are causing these sex differences. Our lab has demonstrated cell-intrinsic sex differences with respect to growth, clonogenicity, and in vivo tumorigenesis using an isogenic murine astrocyte cell culture model of GBM (Nf1 -/-; DNp53). Male astrocytes are more likely to adapt and grow when exposed to irradiation-induced stress compared to female astrocytes, which are more likely to undergo growth arrest after the same exposures. These sex differences in response to treatment can be explained by the different chromatin and transcriptional profiles established from the moment of fertilization that can result in different levels of transcriptional heterogeneity, a well-established cause for resistance to induced stress in GBM. However, so far sex has not been considered when exploring transcriptional heterogeneity. As such, we used 10x Genomics technology to extract single nuclei RNA (snRNA) and ATAC (snATAC) sequencing from the same nuclei and identified different levels of transcriptional heterogeneity in male and female astrocytes cells from our model. We also identified different chromatin profiles of crucial genes, such as Sox2, an important TF for the reprogramming of differentiated GBM into stem-like cells, which can then form tumors in vivo. We expect for the male astrocytes to be more effective in escaping growth arrest and adapting to irradiation-induced stress because of these different transcriptional and chromatin profiles. Our work will identify epigenetic mechanisms by which male cells are more able to adapt to treatment-induced stress compared to female cells and will further advance our understanding of sex differences in treatment-response in GBM and cancer overall, while giving avenues for the development of novel treatments.
INNV-17. EXTRANEURAL METASTASES OF MYXOPAPILLARY EPENDYMOMA TREATED WITH DOSE-DENSE TEMOZOLOMIDE AND LAPATINIB
Authors: Devon Riegel, Jennifer Connelly
Journal: Neuro-Oncology (2022)

Abstract Myxopapillary ependymomas (MPEs) are recently reclassified WHO grade II tumors that usually affect the sacral spinal cord. MPEs have a high recurrence rate, especially if gross total resection (GTR) is not achieved. Recurrence tends to be local, but distant CNS metastases are not uncommon. Rarely, MPEs can metastasize to extraneural locations, including the lungs. Patient outcomes are worse with recurrence, and there is an absence of consensus for the best treatment approach for extraneural MPE metastases. The standard of care for initial MPE diagnosis is GTR, or subtotal resection (STR) with radiotherapy. Systemic treatment has shown inconclusive results regarding improvement of MPE outcomes. A recent phase II clinical trial demonstrated the efficacy of using dose-dense temozolomide (TMZ) and lapatinib for the treatment of metastatic, recurrent ependymoma. We present a case of a 35-year-old female with a history of MPE status post GTR who developed extraneural metastases 11 years after her initial resection. Sites of metastases included multiple bilateral intrapulmonary and pleural based masses with pleural effusion and pelvic mass. Her original site of disease had no evidence of recurrence and imaging of the neuro-axis was negative. Biopsy of the lung mass and pelvic mass confirmed metastatic MPE. She proceeded with dose-dense TMZ and lapatinib and had a positive radiographic response after 3 cycles of treatment. This is the first known case of extraneural metastases of MPE to demonstrate response to dose-dense TMZ and lapatinib.
EPCO-27. REVEALING TUMOR HETEROGENEITY AND IMMUNE MICROENVIRONMENT IN NF-1 GLIOMA BY SINGLE-CELL GENE EXPRESSION PROFILING
Authors: Fulvio D'Angelo, Luciano Garofano, Young Taek Oh, Ryan Najac, Michele Ceccarelli, Marica Eoli, Marc Sanson, Anna Lasorella, Antonio Iavarone
Journal: Neuro-Oncology (2022)

Abstract Germline mutations in the NF1 tumor suppressor gene cause neurofibromatosis type 1 (NF-1), the most common human genetic cancer predisposition syndrome. NF1 patients are prone to develop peripheral and central nervous system tumors, with glioma occurring in 15-20% of patients. The molecular profiles of glioma from patients with NF-1 have been previously explored by large scale DNA and RNA bulk analyses, providing the first comprehensive genetic landscape of NF-1 glioma. However, the intra-tumoral architecture and the microenvironment organization remained largely concealed within the global bulk profile. Here, to characterize the complexity of molecular states of both malignant and non-malignant cells, we profiled NF-1 glioma gene expression at individual cell resolution. We performed single-nuclei RNA sequencing (Nucseq) on frozen NF-1 glioma specimens, including high-grade and low-grade tumors. The functional classification of the transcriptional programs activated in the individual tumor cells identified three main molecular states that partially overlapped with the cellular subtypes of sporadic glioblastoma. Tumor cells of low-grade NF-1 glioma exhibited marked enrichment of neuronal functions, suggesting that the gliomagenesis process in the context of NF1 inactivation might be supported by specialized neuronal activities. Conversely, the proliferative state characterized high-grade NF-1 glioma, indicating that cell cycle-associated pathways are drivers of tumor progression. The single-cell deconvolution of immune microenvironment in NF-1 low grade glioma revealed infiltration of CD8+ T lymphocytes and the active crosstalk of these cells with pro-inflammatory macrophages and mature dendritic cells, supporting a possible anti-tumor cell-mediated response. The identification of divergent tumor cell states as well as the characterization of the immune infiltrates provide new insights for accurate stratification of NF-1 glioma patients and to inform personalized therapeutics.
EPCO-01. EPIGENETIC REPROGRAMMING SHAPES THE MOLECULAR AND CELLULAR LANDSCAPE OF SCHWANNOMA
Authors: S John Liu, Tim Casey-Clyde, Jason Swinderman, Nam Woo Cho, Harish Vasudevan, Kyla Foster, Melike Pekmezci, William Chen, Javier Villanueva-Meyer, Kamir J Hiam-Galvez, Danielle Swaney, Abrar Choudhury, Jonathan Breshears, Erica Stevenson, Kuei-Ho Chen, Brian Lien, David Wu, Ursula Lang, Stephen Magill, Daniel Lim, Michael McDermott, Mitchel S Berger, Arie Perry, Nevan J Krogan, Matthew Spitzer, Luke Gilbert, Philip Theodospoulos, David Raleigh
Journal: Neuro-Oncology (2022)

Abstract DNA methylation profiling provides robust classification of nervous system tumors, but mechanisms driving epigenetic identity of individual tumor types are incompletely understood. Integrating DNA methylation profiling (n=76), RNA sequencing (n=24), single-cell RNA-sequencing (n=9), and mass cytometry (n=9), we discovered vestibular schwannomas are comprised of two epigenetic groups distinguished by neural crest development pathways or repair and regeneration pathways driving immune infiltration. Analyses of preoperative magnetic resonance imaging studies (n=66) or paired primary and recurrent schwannomas (n=13) suggested radiotherapy was sufficient but not necessary for epigenetic reprogramming of neural crest enriched schwannomas into immune enriched schwannomas. In support of this hypothesis, DNA methylation profiling, RNA sequencing, single-cell RNA sequencing, proteomic mass spectrometry, and lymphocyte migration assays demonstrated radiotherapy epigenetically reprogramed viable schwannoma cells to secrete immunomodulatory signals and recruit lymphocytes in vitro. Genome-wide CRISPRi screens identified histone acetyltransferases or DNA methyltransferases driving schwannoma radiotherapy responses, including the epigenetic regulators KDM5C or KDM1A. CRISPRi and lymphocyte migration assays ± radiotherapy confirmed KDM5C drives schwannoma immune infiltration whereas KDM1A inhibits schwannoma immune infiltration. To define genomic mechanisms underlying epigenetic group identity, we performed pooled CRISPRi screening coupled with single-cell RNA sequencing (Perturb-seq) of 44 schwannoma markers. In parallel, we developed single nuclei profiling of chromatin accessibility through paired ATAC sequencing and RNA sequencing coupled with pooled CRISPRi screening (snARC-seq) of 54 epigenetic regulators identified by our genome-wide CRISPRi screen. Functional genomic approaches revealed the tyrosine phosphatase PTPRG as a regulator of survival, and KDM5C and KDM1A as regulators of inflammation. In summary, we report two epigenetic groups of schwannomas and mechanisms underlying epigenetic group identity using a new functional genomic technique allowing for simultaneous interrogation of single-cell epigenetic and gene expression changes in the context of genetic and therapeutic perturbations. These data elucidate a novel epigenetic mechanism of action of radiotherapy.
ANGI-01. METASTATIC ESTHESIONEUROBLASTOMA RECURRENCE AFTER 19 YEARS OF REMISSION: A SYSTEMATIC REVIEW WITH CASE ILLUSTRATION
Authors: Khashayar Mozaffari, Anjali Pradhan, Kunal Patel, Andrew Vivas, Isaac Yang
Journal: Neuro-Oncology (2022)

Abstract INTRODUCTION Esthesioneuroblastoma (ENB) is a rare malignant neoplasm of the olfactory epithelium with an estimated incidence of 0.4/million. It can directly extend along the cribriform plate in order to metastasize to the central nervous system. However, non-contiguous intracranial involvement without recurrence at the primary site is extremely uncommon. In this report, the authors review the literature and present a case of non-contiguous intracranial metastasis of ENB without recurrence at the primary site. To the best of our knowledge, this case presents the longest disease-free interval reported in the literature. METHODS A systematic review of literature was conducted in accordance with the PRISMA guidelines. Additionally, the presentation, surgical management, and post-operative outcomes of an 82-year-old female with non-contiguous intracranial metastasis of ENB after 19 years of remission are described. RESULTS A total of 137 deduplicated works were identified after the search. Of these, 6 papers satisfied our inclusion criteria for our systematic review. Average age at presentation was 50.8 years (range: 26-66) and 52.6% of patients were female. A majority of cases achieved gross-total resection and received adjuvant radiotherapy for initial treatment. The median interval to intracranial metastasis was 6 years from the time of primary tumor presentation. The median overall survival from ENB recurrence with non-contiguous intracranial metastasis was 12 months. CONCLUSIONS ENB is a highly recurrent tumor and harbors the potential to involve the intracranial space even years after remission. Intracranial involvement entails poor overall survival. Lifetime radiographic follow-up should be considered in all patients with ENB.
EXTH-37. TARGETING THE IDH1R132H MUTATION IN GLIOMAS BY CRISPR-CAS PRECISION BASE EDITING
Authors: Remi Weber, Flavio Vasella, Artsiom Klimko, Manuela Silginer, Martine L M Lamfers, Luca Regli, Gerald Schwank, Michael Weller
Journal: Neuro-Oncology (2022)

Abstract Gliomas are the most frequent malignant primary brain tumors and continue to lack curative therapies. To aid the development of novel therapeutic approaches, a better understanding of glioma-specific molecular patterns is essential. We therefore investigate the role of the canonical point mutation in the isocitrate dehydrogenase 1 gene (IDH1R132H). Its early occurrence in oncogenesis and ubiquitous expression suggests a causative driver function; however, current literature is inconclusive on the impact of IDH1R132H, requiring innovative approaches to further elucidate the functional consequences and therapeutic potential of IDH1R132H. METHODS Clustered, regularly interspaced, short palindromic repeat (CRISPR)-Cas precision base editing systems enable the alteration of a specific base pair without suffering from off-target editing associated with previous, double strand brake-inducing systems and thus making them an ideal tool to revert the point mutation in IDH1R132H gliomas. RESULTS By selecting a CRISPR-Cas precision base editing system, the IDH1R132H site was edited with editing efficiencies of up to 50%. Primary patient derived cell lines and organoids were corrected in their IDH1R132H locus. Phenotypical changes, such as a change in IDH1R132H protein levels, 2-hydroxyglutarate concentration as well as proliferation rates were observed upon the reversal of the point mutation. Furthermore, the precision base editing system was packaged into a dual-AAV-vector split intein system and showed successful in vitro gene editing. In conclusion, this precise genetic intervention provides a methodology to create accurate patient derived models to analyze the impact of IDH1R132H on glioma biology and provides a framework for in vivo gene therapy.
PATH-14. MOLECULAR ANALYSIS OF ASTROCYTOMAS WITH A DOMINANT PILOMYXOID MORPHOLOGY
Authors: Samantha Ritacco, Brennan Decker, J Keith Killian, Erik Williams, Mirna Lechpammer
Journal: Neuro-Oncology (2022)

Abstract Pilomyxoid astrocytomas (PMAs) are tumors of infancy occurring in the hypothalamic chiasmatic region, with somewhat worse prognosis than pilocytic astrocytomas (PAs). We report molecular findings in PAs with a dominant pilomyxoid histomorphology across the various age groups and CNS locations. Foundation Medicine comprehensive genomic profiling (CGP) (FoundationOne, FoundationOneCDx, FoundationOneHeme) were performed during routine clinical care. A retrospective cohort analysis was done to evaluate the genomic characteristics of PMAs. 57 PMAs were identified, of which 6 occurred in patients &amp;lt; 12 months old (10.5%); 14 in 1-3 year old (24.6%); 15 in 4-10 y.o. (26.3%); 12 in 11-20 y.o. (21.1%), and 10 occurred in young adults 21-39 y.o. (17.5%). 30/57 (52.6%) were female and 27/57 (47.4%) male. 55 PMAs occurred in the brain and 2 in the spinal cord. Tumors were categorized by notable rearrangements and point mutations. The common fusion, KIAA1549:BRAF, was detected in 23/57 samples (40.4%). BRAF fusions with other gene partners (BRAF, TAX1BP1) occurred in 2, and BRAF V600E mutations in 4 patients. An additional 4 patients had FGFR1 fusions (FGFR1, TACC1) and 14 had FGFR mutations. Other alterations included 3 NF1 mutations, 2 NTRK family receptor tyrosine kinase fusions, 2 RAF gene fusions, 2 other (ATRX, ALK) alterations, and 1 VUS. Tumor Mutational Burden was under 10 mut/Mb for all patients. In this cohort of 57 PMAs, MAPK pathway genetic alterations were dominant drivers, similar to other PAs. We also observed other frequent genetic alterations not involving BRAF, including FGFR and NTRK gene family members that have FDA approved targeted inhibitors. While these also likely cause MAPK pathway activation, these non-BRAF variants are less common in PAs. Our report contributes to growing evidence that PMAs harbor a distinct molecular alteration profile, which is becoming increasingly important in the context of the evolving targeted therapeutics landscape.
EPCO-33. DIFFERENTIAL EXPRESSION OF AN ENDOGENOUS RETROVIRAL ELEMENT [HERV-K(HML-6)] IS ASSOCIATED WITH REDUCED SURVIVAL IN GLIOBLASTOMA PATIENTS
Authors: Ashish Shah, Vaidya Govindarajan, Tara Doucet-O'Hare, Sarah Rivas, Leo Ampie, Catherine DeMarino, Yeshavanath Banasavadi-Siddegowda, Yong Zhang, Kory Johnson, Fahad Almsned, Mark Gilbert, John Heiss, Avindra Nath
Journal: Neuro-Oncology (2022)

Abstract INTRODUCTION Comprising approximately 8% of our genome, Human Endogenous RetroViruses (HERVs) represent a class of germline retroviral infections that are regulated through epigenetic modifications. In cancer cells, which often have epigenetic dysregulation, HERVs have been implicated as potential oncogenic drivers. However, their role in gliomas is not known.Given the link between HERV expression in cancer cell lines and the distinct epigenetic dysregulation in gliomas, we utilized a tailored bioinformatic pipeline to characterize and validate the glioma retrotranscriptome and correlate HERV expression with locus-specific epigenetic modifications.Method:A custom workflow was used to quantify HERV expression in our cell lines of interest. Cell-line methylation was quantified using a custom script. We generated primers specific for the Human endogenous Mouse mammary tumor (MMTV)-Like virus 6 (HML-6). Visualization of RNA transcripts was performed using RNA-scope. Clinical data was obtained using the R package, TCGABiolinks. RESULTS The A172 cell line had significantly higher mean overall HERV expression relative to the M059J and H4 cell lines (p&amp;lt; 0.0001 for both). A172 cells had significantly lower mean number of CpG islands relative to M059J cells and H4 cells (p&amp;lt; 0.0001 for both). There was a significant inverse correlation between mean beta value and FC HERV expression (R=-0.57, p=0.01). qPCR confirmed robust expression of the HML-6 locus in cell culture and neurospheres. Elevated ERVK3-1 expression was associated reduced survival among IDHwt GBM patients (18.3 vs. 15.1 months, p=0.039). This was preserved among IDH mutant (IDHm) GBM as well (17.9 months vs. 14.0 months, p=0.0088). CONCLUSION In gliomas, HERV expression correlates with loss of DNA methylation at HERV loci. HML-6 is overexpressed in highly invasive glioblastoma cell lines and patient-derived neurospheres. We have demonstrated a potential survival detriment associated with elevated expression of the HML-6 product, ERVK3-1.